Based on new evidence and a review of prior studies, the current committee did not find any new associations between the relevant exposures and immune outcomes that warranted a change in level of evidence of association. Current evidence supports the findings of earlier studies that there is inadequate or insufficient evidence to determine whether there is an association between exposure to the chemicals of interest (COIs) and any specific diseases involving immune suppression, allergy, autoimmunity, or inflammation.
Immune-system disorders affect more than 23.5 million Americans (NIEHS, 2012), with some sources estimating that as many as 50 million Americans are currently affected by immune-system disorders (AARDA, 2018). The causal factors for immune-system disorders are mainly unknown; however, it has been hypothesized that they most likely reflect both genetic and environmental factors. The immune system plays three important roles in the body:
- It defends the body against infectious pathogens, including viruses, bacteria, and other disease-producing microorganisms.
- It helps defend against cancer by destroying mutated cells that may otherwise develop into tumors and providing immunity against tumors.
- It provides resident immune cells that are specially adapted for different tissues and organs (such as microglia in the central nervous system and Kupffer cells in the liver) to help regulate the functional activity and integrity of those tissues.
This chapter begins with an overview of the various types of health problems that can arise as a result of a malfunction of the human immune system, such as immune suppression, allergic diseases, autoimmune diseases, and inflammatory diseases. Outcomes related to infectious agents would be included in this chapter, but no studies had specific enough information on exposure to warrant inclusion. Following the brief description of the types of immune dysfunction, the findings from previously reviewed literature and the conclusions from prior updates are summarized regarding the epidemiologic evidence concerning an association between exposure to any one of the COIs, that is, 2,4-dichlorophenoxyacetic acid (2,4-D); 2,4,5-trichlorophenoxyacetic acid (2,4,5-T); picloram; dimethylarsinic acid (DMA or cacodylic acid); and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). As described in Chapter 3, studies of exposure to polychlorinated biphenyls (PCBs) and other dioxin-like chemicals were also considered informative if their results were reported in terms of TCDD toxic equivalents (TEQs) or as concentrations of specific congeners of dioxin-like chemicals. Studies that report TEQs based only on mono-ortho PCBs (which are PCBs 105, 114, 118, 123, 156, 157, 167, and 189) are considered even though their TEQs are several orders of magnitude lower than those of the non-ortho PCBs (77, 81, 126, and 169), based on the revised World Health Organization (WHO) toxicity equivalency factor (TEF) scheme of 2005 (La Rocca et al., 2008; van den Berg et al., 2006). Epidemiologic findings from five newly identified studies are reviewed. Biologic plausibility data on the effects of the COIs on the immune system are summarized. The chapter ends with a synthesis of the studies and what they contribute to the evidence base and then the committee’s conclusion regarding the association of exposure to the COIs and effects on the immune system. The studies reviewed in this chapter are limited to those investigating effects on the immune system from exposures that occurred to adults. Immune effects stemming from perinatal exposures are discussed in Chapter 8. Table 1, which can be found at www.nap.edu/catalog/25137, summarizes the results of studies reviewed in the VAO series related to immune disorders.
There are four major categories of immune dysfunction, which are not mutually exclusive: immune suppression, allergy, autoimmunity, and inflammatory dysfunction (inappropriate or misdirected inflammation). Most times, immune suppression manifests itself as an increased incidence of infections or an increased risk of neoplasia. Allergic, autoimmune, and inflammatory disorders can be manifested as diseases that affect virtually any tissue. It is often difficult to diagnose such diseases, so they may not always be medically categorized as immune disorders.
The suppression of immune responses can reduce resistance to infectious disease and increase the risk of cancer. Infection with the human immunodeficiency virus (HIV) is a well-recognized example of an acquired immune deficiency in which a specific type of lymphocyte (CD4+ T cell) is the target of the virus (Okoye and Picker, 2013). The decline in the number of CD4+ T cells after HIV infection correlates with an increased incidence of infectious diseases, including fatal opportunistic infections, and with an increased incidence of several types of cancer (Davis, 1998). The treatment of the cancer with toxic chemotherapeutic drugs suppresses the immune system by inhibiting the generation of new white blood cells by the bone marrow and blocking proliferation of lymphocytes during an immune response. Both of those examples represent severe immune suppression in which the adverse outcome is easily detected with clinical measurements.
Immune suppression can also result from exposure to chemicals in the workplace or in the environment and can manifest as recurrent infections, opportunistic infections, a higher incidence of a specific category of infections, or a higher incidence of many forms of cancer (Saberi Hosnijeh et al., 2013; Warren et al., 2000). However, unless the immune suppression is severe (as occurs in rare cases of genetic disorders of immunity), it is often difficult to obtain clinical evidence that directly links chemically induced changes in immune function to increases in infectious diseases or cancers because many confounding factors and effect modifiers can influence a person’s ability to combat infection. These factors include age, vaccination status, the virulence of the pathogen, the presence of other diseases (such as diabetes), stress, smoking, and the use of drugs or alcohol. Therefore, immunotoxicology studies are often conducted in laboratory animals to understand the scope and mechanism of chemical-induced immune suppression. The results of such studies can be used to develop biomarkers to assess the effects in human populations. Infectious disease models in animals can also be used to determine whether the pattern of disease changes with chemical exposure.
The immune system sometimes responds to a foreign substance that is not pathogenic; such immunogenic substances are called allergens. Like most immune-based diseases, allergic diseases have both environmental and genetic risk factors. Their prevalence has increased in many countries in recent decades (Linneberg et al., 2000; Simpson et al., 2008; Sly, 1999). The major forms of allergic diseases are asthma, allergic rhinitis, atopic dermatitis, and gastrointestinal responses. In immediate hypersensitivity, the response to some allergens, such as pollen and bee venom, results in the production of immunoglobulin E (IgE) antibodies. Once produced, IgE antibodies bind to mast cells, which are
specialized cells that are found in tissues throughout the body, including the lungs, the intestinal wall, and blood-vessel walls. When a person is exposed once again to the allergen, it binds to the antibodies on the mast cells and causes them to release histamine and leukotrienes, which produce the symptoms associated with an allergic response. In delayed-type hypersensitivity reactions, also known as cell-mediated immunity, other allergens, such as poison ivy and nickel, activate allergen-specific lymphocytes (memory T cells) at the site of contact (usually the skin) that release substances that cause inflammation and tissue damage. Some allergic responses, such as those to food allergens, may involve a combination of allergen-specific lymphocyte-driven and IgE-driven inflammation. Allergic responses may manifest in specific tissues (such as the skin, eyes, airways, and gastrointestinal tract) or may result in a system-wide response called anaphylaxis.
The National Institutes of Health’s Autoimmune Diseases Coordinating Committee recognizes 80 different autoimmune diseases and conditions that affect the cardiovascular, respiratory, nervous, endocrine, skin, gastrointestinal, hepatic, and excretory systems (NIH Autoimmune Diseases Coordinating Committee, 2005). These diseases affect both men and women, but most of them affect more women than men (Fairweather et al., 2008). Genetic predisposition, age, hormone status, and environmental factors, such as the presence of infectious diseases and stress, are known to affect the risk of developing autoimmune diseases. Different autoimmune diseases can occur in the same person and tend to cluster in families. The development of one autoimmune condition is also a risk factor for the development of other immune-related diseases and for some types of cancer (Landgren et al., 2010).
Autoimmunity occurs when an individual’s immune system fails to recognize self and attacks tissues as though they were foreign. Inappropriate immune responses that cause autoimmunity originate with either cell-mediated or humoral-mediated immune systems and can be directed against a wide variety of tissues or organs. For example, the autoimmune reaction in multiple sclerosis targets the myelin sheath of nerve axons; in Crohn’s disease, the intestinal epithelium; in type 1 diabetes mellitus, the insulin-producing islet cells of the pancreas; and in rheumatoid arthritis, the joint synovium and other proteins associated with connective tissue.
Other systemic autoimmune diseases also occur. Systemic lupus erythematosus is an autoimmune disease in which multiple organs are targeted by a variety of autoantibodies. Patients display a variety of non-specific signs and symptoms such as joint pain or fatigue that makes timely diagnosis challenging. A characteristic rash across the cheeks and nose and a sensitivity to sunlight are common symptoms, but oral ulcers, arthritis, pleurisy, proteinuria, and neurologic signs may also be present. Almost all of those affected with systemic lupus erythematosus test positive for antinuclear antibodies, specifically antibodies directed at
double-stranded DNA. The cause of systemic lupus erythematosus is unknown, but environmental and genetic factors have been implicated. The environmental factors that are thought to trigger it include infections, antibiotics (especially those in the sulfa and penicillin groups) and some other drugs, ultraviolet radiation, extreme stress, and hormones (Kamen, 2014). Occupational exposures to such chemicals as crystalline silica, solvents, and pesticides have also been associated with systemic lupus erythematosus (Cooper and Parks, 2004; Parks and Cooper, 2005).
Inflammatory diseases (also referred to as auto-inflammatory diseases) make up a more recently identified category of immune-related disorders and are characterized by exaggerated, excessively prolonged, or misdirected dysfunctional inflammatory responses (usually involving immune cells). Tissue disease can result from this inappropriate inflammation, which can affect virtually any organ. Examples of the diseases and other conditions that are most often included in other disease categories but that are also considered to be inflammatory diseases are: coronary artery disease, asthma, eczema, chronic sinusitis, hepatic steatosis, psoriasis, celiac disease, and prostatitis. Inflammatory diseases often co-occur with one another, which has resulted in the categorizing of different but linked inflammatory diseases together as a single chronic inflammatory disorder (Borensztajn et al., 2011). Ordinarily, inflammation is advantageous in fighting infection. It is one component of the normal host response to infection and is mediated by innate and adaptive immunity. Innate inflammatory responses involve the rapid mobilization of macrophages, granulocytes, and natural killer cells to the area of infection, where they produce toxic metabolites that kill pathogens. The adaptive immune response follows with specific antibodies and cell-mediated immunity that add to the inflammatory process. Interactions among innate immune cells and epithelial and endothelial cells are important in regulating the magnitude of inflammation, and improperly regulated inflammation can contribute to diseases that arise in non-lymphoid tissues, such as the lungs, skin, nervous system, endocrine system, and reproductive system.
Inappropriate inflammation also appears to play a role in promoting the growth of neoplasms (Bornschein et al., 2010; Hillegass et al., 2010; Landgren et al., 2010; Porta et al., 2011; Winans et al., 2010); examples can be seen in the higher prevalence of specific cancers in patients who have such inflammatory diseases as inflammatory bowel disease (Lucas et al., 2010; Viennot et al., 2009; Westbrook et al., 2010), prostatitis (Sandhu, 2008; W. Wang et al., 2009), and psoriasis (Ji et al., 2009).
The studies reviewed in this section are limited to effects on the immune system due to exposures that occurred to adults. Immune effects stemming from
perinatal exposures are discussed in Chapter 8: Reproductive Health Effects and Effects on Descendants.
Two studies of Vietnam veterans reported a statistically significant difference of single immune measures of veterans exposed to Agent Orange compared with veterans without diseases and with age-matched healthy controls. Among Air Force Health Study (AFHS) veterans, Ranch Hands with the highest TCDD levels had statistically significantly elevated absolute counts of CD20 cells, the sum of natural killer (NK) cell antigens (CD16 and CD56) and total CD3+ cells compared with the AFHS controls (Michalek et al., 1999b). H. A. Kim et al. (2003) conducted an immunotoxicologic study of Korean Vietnam veterans who had been deployed to areas that were known to be sprayed with Agent Orange and found elevated levels of IgE and IgG1 among exposed veterans compared with age-matched healthy controls. Other studies of Vietnam veterans and populations exposed to the COIs did not find similar elevations for these immune markers. Thus, there were no consistent findings indicative of immunosuppression, an increased risk of autoimmunity (usually as measured with autoantibodies), or biomarkers of atopy or allergy (such as increased IgE concentrations). Much of the focus of the studies was on measuring CD4+:CD8+ T-cell ratios (T4:T8). The T4:T8 ratio is an effective biomarker of the progression of HIV-induced AIDS (acquired immune deficiency syndrome), but the TCDD-exposure animal data indicate that it is not an immunologic index that is expected to be altered. The results of a survey of Australian Vietnam veterans (O’Toole et al., 2009) showed purportedly significant increases in the prevalence of a number of conditions in which immune function may play a prominent role, but the study’s methods were deemed unreliable.
The occupational exposure studies evaluated by VAO committees have examined the concentrations of lymphoid populations in circulation, such as CD4+ T cells, CD8+ T cells (and their ratio), and NK cells; cell-mediated immunity (the delayed-hypersensitivity response); serum concentrations of immunoglobulins, such as IgM, IgG, and IgA; concentrations of complement, such as C3 and C4; and concentrations of cytokines, such as IL-1, IL-2, interferon-γ, IL-4, IL-6, and tumor necrosis factor (TNF)-α. A few studies also included disease or condition endpoints, such as rheumatoid arthritis, systemic lupus erythematosus, immune suppression, and sensitivity to fungal infection. Ex vivo analyses included measures of NK activity, lymphoid mitogen-induced proliferation, and the mixed lymphocyte response against allogeneic cells. Some studies identified one or more dioxin-related shifts in immune measures, but many reported no significant differences in the same measures. Saberi Hosnijeh et al. (2012a) reported a positive correlation between plasma TCDD concentrations and decreased levels of cytokines, chemokines, and growth factors in Dutch workers who produced and formulated chlorophenoxy herbicides. In additional studies, Saberi Hosnijeh et al. (2012b, 2013a) assessed TCDD levels with respect to several other immunological parameters and found no differences in hematologic
measurements other than an increase in the T4:T8 ratio (p = 0.05) when high- and low-exposure groups were compared. In the third study, Saberi Hosnijeh et al. (2013a) reported on the levels of interleukin 1 receptor antagonist (IL-1RA) as well as of the soluble forms of CD27 and CD30, immunomodulatory members of the TNF receptor superfamily. In this case they found no association of TCDD level with CD27 or CD30; however, IL-1RA was significantly decreased in those with higher TCDD levels after adjusting for concurrent chronic disease. This result is also consistent with a degree of immune system impairment being associated with high exposure to TCDD. Similarly, the occupational exposure studies that examined NK concentrations reported the full spectrum of results: no alterations (Halperin et al., 1998), a decrease (Faustini et al., 1996), and even an increase in NK numbers (Jennings et al., 1988) in different populations of people exposed to dioxin.
Several environmental exposure studies have been published, with inconsistent findings. Some studies reported alterations in immune measures associated with dioxin exposure. For example, Van den Heuvel et al. (2002) reported a negative correlation between increased serum TEQ levels and eosinophil counts, NK-cell counts, and levels of cat dander, house dust mite, and grass pollen IgEs (measured by radioallergosorbent tests); but a positive correlation with IgA levels. These alterations, however, were not seen consistently in other studies. Baccarelli et al. (2002) found no changes in IgA levels but saw changes in IgG levels in the Seveso population. Svensson et al. (1994) found that NK-cell numbers were reduced with increasing concentrations of persistent organic chemicals, but Lovik et al. (1996) found no difference in NK numbers or activity.
Some early studies of the Quail Run Mobile Home Park population who were exposed to TCDD-contaminated soil reported that dioxin exposure was associated with a reduction in a specific type of cell-mediated immune response, the delayed type hypersensitivity response (Andrews et al., 1986; Hoffman et al., 1986; Knutsen et al., 1987; Stehr-Green et al., 1987). However, several studies of the Times Beach population, another dioxin-exposed population, did not find any alteration of the delayed type hypersensitivity response (Knutsen, 1984; Stehr et al., 1986; Webb et al., 1987). An analysis of National Health and Nutrition Examination Survey (NHANES) data found that exposure to dioxin-like PCBs was associated with an increase in self-reported arthritis (D. H. Lee et al., 2007a), but De Roos et al. (2005b) did not find this association. Spector et al. (2014) assessed immune function in 109 postmenopausal women who participated in a year-long study of exercise and health in Seattle. At baseline, the concentrations of mono-ortho PCBs 105, 118, and 156 were not associated with changes in lymphocyte proliferation assays; after 1 year, however, a decrease in lymphocyte proliferation was associated with increased levels of this group of PCBs (p = 0.039).
Nakamoto et al. (2013) gathered fasting blood samples to assess environmental exposure to dioxin-like polychlorinated dibenzo-p-dioxins (PCDDs), polychlorinated dibenzofurans (PCDFs), and PCBs and found an association of
exposure to dioxins with differential leukocytes and a significantly decreased incidence of reported atopic dermatitis. Gallagher et al. (2013) examined the link between environmental exposures to the COIs and autoimmunity in women. Among women, after adjustments for mercury blood level, race, menopausal status, diet, and body mass index (BMI), the total TEQs for PCBs were significantly associated with antinuclear antibody positivity (intensity ≥ 3) for each higher quartile compared with the lowest and as an overall trend (p < 0.001). After adjusting for mercury blood level, race, menopausal status, diet, and BMI, total TEQs for PCBs were significantly associated with antinuclear antibody positivity for each quartile compared with the referent group, and the overall trend was significant (p < 0.001). Turunen et al. (2012) derived the total toxic equivalence for 17 PCDD/F and 37 PCB congeners in blood samples from 123 men and 132 women from a population with high fish consumption and analyzed their relationship with C-reactive protein, an indicator of inflammation. No evidence of a trend across the tertiles of overall TEQ concentration was seen for either the men or the women.
Based on these mixed results, the variety of endpoints assessed, and the poor methods used in some studies, VAO committee have concluded that epidemiologic data were either insufficient or inadequate to determine an association between exposure to the COIs and immunosuppression, allergic disease, or autoimmune disease.
Since Update 2014, five new published studies have been identified: one focused on New Zealand Vietnam veterans, three among occupational cohorts, and one on environmental exposures in Vietnam.
Cox et al. (2015) used hospital discharge records from 1988 to 2009 to identify prevalent health conditions in 2,783 male New Zealand veterans who served in Vietnam during 1964 to 1972. Age-specific hospitalization rates were calculated using the total number of annual hospitalizations published by the Ministry of Health and the average annual resident population. Standardized hospitalization rates and 99% confidence intervals (CIs) were calculated for the veteran cohort and the general population for several noncancerous conditions and a standardized hospitalization ratio (SHR) was calculated using the two rates. Results were presented for three categories of arthritis: rheumatoid arthritis, infective arthritis, and osteoarthrosis. A statistically significant increased risk was observed for rheumatoid arthritis (n = 25; SHR = 1.70, 99% CI 1.03–2.36) and osteoarthrosis (n = 179; SHR = 1.32, 99% CI 1.12–1.51) but not for infective arthritis (n = 26; SHR = 1.39, 99% CI 0.85–1.92). However, arthritis is not usually
a condition that requires hospitalization, so this study likely underestimates the prevalence or detects only persons with an unusually complicated disease process. In addition, hospitalization for systemic lupus erythematosus was not statistically significantly increased among veterans (n = 5; SHR = 1.82, 99% CI 0.22–3.41). Exposure to the COIs was not validated through serum measurements, and the study did not control for smoking or ethnicity or other potentially important risk factors.
Parks et al. (2016) used data collected from the prospective, longitudinal Agricultural Health Study (AHS) to examine the associations between rheumatoid arthritis and pesticide exposures among the spouses of licensed pesticide applicators from Iowa and North Carolina who were part of the AHS. Specific pesticide exposures included exposure to herbicides 2,4-D and dicamba and to several insecticides and fungicides; only 2,4-D and dicamba are considered relevant to committee’s charge. Details of the AHS study design and data collections are found in Chapter 5. Cases of rheumatoid arthritis were identified based on self-report. Cases in which disease-modifying antirheumatic drugs or other medications for rheumatoid arthritis were identified were considered to be probable instances of rheumatoid arthritis, and those individuals for whom a medical record review allowed confirmation of the diagnosis were considered to be definite cases of rheumatoid arthritis. Surveys collected information on age, tobacco use, menopausal status, childhood farm exposure, non-specific household and specific farm pesticide exposure, farm activities related to pesticides and other activities such as sun exposure, exposure to solvents and exposure to pesticides through mixing or application. A total of 275 cases of rheumatoid arthritis were identified among the 24,018 women participants, of which 132 were incident cases (that is, they developed after the start of the study, having been reported in phase II or III surveys but not phase I). Cases reported in phase I were considered prevalent, that is, as having developed before the start of the study. After adjusting for age, state of residence, and smoking history, the researchers found a statistically significant association between all-cases rheumatoid arthritis (incident and prevalent) and any specific pesticide exposure (odds ratio [OR] = 1.4; 95% CI 1.0–1.8; p < 0.05). Incident cases of rheumatoid arthritis were not associated with any exposure (OR = 1.4; 95% CI 0.93–2.1). A decreased association was observed for rheumatoid arthritis and exposure to 2,4-D with all cases (OR = 0.75; 95% CI 0.51–1.1) and incident cases (OR = 0.69; 95% CI 0.39–1.2). Likewise, a decreased association was observed for rheumatoid arthritis and exposure to dicamba based on 7 cases (OR = 0.68; 95% CI 0.32–1.5). The effect estimate was not presented for incident cases of rheumatoid arthritis and dicamba exposure because there were fewer than 5 cases. The only individual pesticide to demonstrate a statistically significant association with rheumatoid arthritis was glyphosate (OR = 1.4; 95%
CI 1.0–2.1; p < 0.05). Thus, this study does not support an association of 2,4-D in the development of rheumatoid arthritis. The strengths of the study are the sample size, duration of follow-up, and specific pesticide data. The self-reported nature of the exposure without quantification could lead to under- or over-estimation as well as a misclassification of the agents of exposure. Para-occupational exposure and agricultural drift were not considered. Though the statistical methods for handling missing data were sound, there was a variety of missing data.
't Mannetje et al. (2018) conducted a morbidity survey among a subset of workers who were employed at the New Plymouth, New Zealand, phenoxy herbicide production plant for at least 1 month between 1969 and 1984. The plant produced 2,4,5-T, and workers were potentially exposed to 2,4,5-T, intermediates of trichlorophenol and other chlorophenols, and TCDD. Workers had previously been recruited and examined as part of the international cohort of producers of phenoxy herbicides led by the International Agency for Research on Cancer (IARC; Kogevinas et al., 1997); see Chapter 5 for more details on the IARC cohort and the New Zealand phenoxy producers. The 't Mannetje et al. (2018) study extended the follow-up period of these workers to approximately 30 years from their last 2,4,5-T production exposure. From the original cohort of 1,025 workers, 631 were living, had a current address in New Zealand, and were below 80 years of age on January 1, 2006. For the current follow-up, 430 of the 631 workers were randomly selected and invited to participate in the morbidity survey, of which 245 (57%) participated. The survey, which was administered in 2007–2008 by face-to-face interview, collected information on demographic factors and health information, including doctor-diagnosed conditions and year of diagnosis. A blood sample was also collected at that time to be analyzed for TCDD, lipids, thyroid hormones, and other parameters. For 111 of the participants, a neurological examination was also conducted. Associations between exposure and health outcomes were assessed using logistic regression models that controlled for age, gender, smoking, BMI, and ethnicity using two methods: (1) working in a TCDD-exposed job (based on occupational records), and (2) serum TCDD concentration ≥10 pg/g lipid (18%). Mean TCDD concentrations were 19 pg/g lipid in the 60 men directly involved in phenoxy or trichlorophenol production and 6 pg/g lipid in 141 men and 43 women who worked in other parts of the plant. Compared with the 124 people in the non-highly exposed jobs, the 121 people who had ever worked in a highly exposed job were no more likely to have doctor-diagnosed nasal allergies, including hay fever (n = 21; OR = 1.00; 95% CI 0.48–2.08). When compared by serum TCDD concentration ≥10 pg/g lipid, no difference in nasal allergies was found (n = 7; OR = 0.80; 95% CI 0.27–2.37).
Cappelletti et al. (2016) performed a retrospective study of 331 male electric arc foundry workers at a single plant in Trentino, Italy, to determine if they had experienced excess mortality from all causes or were at an increased risk for several other diseases, including rheumatoid arthritis, due to occupational exposures to foundry dust. An analysis of the dust found that it contained metals (including
iron, aluminum, zinc, manganese, lead, chromium, nickel, cadmium, mercury, and arsenic), polycyclic aromatic hydrocarbons (PAHs), PCBs, and PCDD/Fs (reported as TEQs). Therefore, the authors could not determine which of the agents were associated with a specific outcome or to what extent. Each of the men had worked at the factory for at least 1 year, and, for the rheumatoid arthritis analysis, they were compared with 32 presumed non-exposed workers (clerks, managers, and watchmen) or the standardized general population of Region Trentino-Alto Adige (where the factory was located) because there were few non-exposed foundry workers and high attrition rates. Company and medical records were used to determine vital status; cause of death was determined from death certificates or other registries. Requests for exemption health care fees were used as a surrogate measure to identify the most prevalent morbid conditions in the general population, which were then applied to the cohort to compute relative risks for each of the conditions. The workers were followed from March 19, 1979 (or their first day of employment), through December 31, 2009, or their date of death. The analysis for rheumatoid arthritis was limited to 235 workers, and effect estimates were calculated using Mantel-Haenszel tests. Although a statistically significant increase of rheumatoid arthritis was found among the workers compared with the age-adjusted provincial population, it was based on three cases, which resulted in an imprecise effect estimate (RR = 6.18, 95% CI 2.00–19.02). This study is most limited by the fact that foundry dust is a complex mixture, which results in an inability to discern the impact of the specific contaminants of the foundry dust on the health outcomes of those exposed workers. Estimates were only adjusted for age group and were not adjusted for other risk factors such as tobacco use, BMI, or other jobs or activities that could also influence health outcomes. The exposure to foundry dust by the general population that was used for comparison was not discussed, although the foundry appears to be in the local vicinity, and emissions from it were reported to be present within a 2-kilometer radius of the foundry.
C. H. Nguyen et al. (2017), studied serum TCDD levels, the expression of AHR, and a variety of pro-inflammatory cytokines in Vietnamese who were either exposed or not exposed to TCDD-like chemicals. The exposed individuals (36 women and 24 men) had lived near the Da Nang Air Base for more than 10 years. The controls were healthy men and women recruited from unsprayed areas in northern Vietnam. Serum levels of TCDD and “other” types of dioxins were measured using a chemical-activated luciferase gene-expression bioassay. RNA was isolated from whole blood for quantitative real-time polymerase chain reaction assays that were used to examine gene expression for AHR, IL-1β, TNFα, IL-6, IL-22, and β-actin. Dioxin levels, presented as bioanalytic equivalents (BEQ)/g fat (1.5 interquartile range), were significantly higher in the TCDD-exposed participants than in the controls—62.03 (40.74–89.42) versus 17.45 (12.05–35.57),
respectively. AHR expression was 16.39 times higher in the exposed versus unexposed controls, which was statistically significant. To determine if there was an association between AHR expression and rheumatoid arthritis in the dioxin-exposed subjects, AHR expression was measured and the 6 people with rheumatoid arthritis were compared to the 54 people without rheumatoid arthritis. While the dioxin-exposed group had higher levels of AHR expression than controls, there were no differences in AHR expression levels between the exposed subjects with and without rheumatoid arthritis. Because of the role they play in the pathogenesis of autoimmune conditions, IL-1β, TNFα, IL-6, and IL-22 were expression levels measured in TCDD-exposed and unexposed populations. The median increase and 1.5-interquartile range for IL-1β was 11.18-fold (6.76–20.72, p < 0.05); for TNFα, 6.64-fold (3.43–13.67, p < 0.05); and for IL-6, 2.93-fold (1.80–5.18, p < 0.05). IL-22 was decreased 0.05-fold (0.01–0.11, p < 0.01). There was, however, no association between cytokine levels and dioxin levels. There was a weak positive correlation between AHR expression and IL-6 and IL-22 expression.
Next the investigators looked at the range of conditions reported in the exposed subjects. Cardiac disease was the most common, affecting 18%. Rheumatologic conditions affected 16.7%, and rheumatoid arthritis affected 10%. The rate for rheumatoid arthritis in the general Vietnamese population was 0.05%. The main strength of this study of Vietnamese individuals living near the Da Nang Air Base is the availability of serum dioxin levels. While the levels of proinflammatory cytokines were significantly elevated in the exposed population, they did not correlate with serum dioxin levels. While there were more cases of rheumatoid arthritis in the exposed population than in the general population, neither the dioxin levels nor pro-inflammatory cytokines were compared between exposed people with and without rheumatoid arthritis. The study demonstrates an increased expression of pro-inflammatory cytokines in persons exposed to dioxins, but there is no information on lifestyle habits, tobacco, obesity, or other rheumatologic disorders or family history that may confound the findings.
Other Identified Studies
Several other studies were identified by the committee but either lacked sufficient exposure specificity or examined biologic markers of effect on the immune system that do not relate to a diagnosable health outcome; these studies were not considered further. Akahane et al. (2017) examined the prevalence of a variety of self-reported conditions, including several connective tissue disorders, in people exposed to PCBs, dioxins (e.g., PCDD/Fs), and dioxin-like chemicals through the ingestion of contaminated rice bran oil following the Yusho accident. Because neither TEQs nor any other quantification of relevant exposures was presented, this study was not considered further.
A study of people who either worked at a transformer and capacitor recycling plant in Dortmund, Germany, or lived in the immediate area and might
have been exposed to dioxin-like (PCBs 105, 114, 118, 156, 157, 167, and 189) and non-dioxin-like PCBs as a result of contamination of the area by the facility confirmed that exposure can modify lymphocyte profiles, but this was not linked with specific health outcomes (Haase et al., 2016). The results are consistent with the literature suggesting that PCB exposure may alter the immune profile, but these data do not provide any evidence of any consistent abnormality. This study did not detect any functional immunodeficiency, and the problems with its design and analysis (detailed in Chapter 5) seriously limit its contribution to the scientific evidence on the effects of exposure to the COIs.
Serdar et al. (2014) used data from the 2003–2004 cycle of NHANES to examine the association of serum, lipid-adjusted PCB levels, and organochlorine pesticide levels (including PCDDs, PCDFs, and dioxin-like PCBs) with hematology and blood chemistry profiles that measured electrolytes, liver and renal function, globulin, total protein, and other factors. Although some statistically significant differences were found between counts and levels in the highest- and lowest-exposed quartiles, all were within the normal ranges and limits for those markers. These measures are indicators, not health outcomes, thus limiting their interpretability concerning immune system conditions.
Given the growing recent understanding of the role of AHR in immune functions (Abe et al., 2014; Biljes et al., 2017; Bock et al., 2017a; Chinen et al., 2015; Huai et al., 2014; Kado et al., 2016; Kimura et al., 2014; Y. H. Lee et al., 2015; Liao et al., 2017; Murray and Perdew, 2016; Stockinger et al., 2014), it is not surprising that there is an extensive body of evidence from experimental studies in cultured cells and animal models indicating that TCDD and other dioxin-like chemicals are immunotoxic to a variety of leukocytes (Kerkvliet, 2009, 2012; Kreitinger et al. 2016). Given that most of the cell types involved in the immune system express the AHR, there are many potential pathways to immunotoxicity. TCDD-induced immunotoxicity is due primarily to changes in adaptive immune responses resulting in the suppression of both antibody-mediated and cell-mediated immunity. Dioxin and other AHR agonists may also reduce the clearance of infections and promote tumor growth through alterations in immune function. TCDD exposure alters macrophages and neutrophils so as to exacerbate some types of inflammation during infections, and it may contribute to the development of chronic inflammatory lung disease (Teske et al., 2005; P. S. Wong et al., 2010). Although there are many examples of dioxin and dioxin-like chemicals having immunosuppressive effects, these chemicals also appear to influence autoimmune diseases, which are viewed as an inappropriate increase in immune function. Therefore, these chemicals may be best described as immunomodulatory.
TCDD has been shown to be a potent immunosuppressive chemical in laboratory animals and cell culture models. Data show that the relative potencies
of TCDD and dioxin-like chemicals on leukocytes are predicted by their TEFs (Frawley et al., 2014; Smialowicz et al., 2008). The exposure of animals to dioxin not only suppresses some adaptive immune responses, but also has been shown to increase the incidence, progression, and severity of various infectious diseases and to increase the development of cancers (Choi et al., 2003; Elizondo et al., 2011; Fiorito et al., 2010, 2011, 2014; Head and Lawrence, 2009; Jin et al., 2010; Sanchez et al., 2010). Also, developmental exposure to TCDD (in utero and via suckling) in mice reduces immune response capacity to the influenza virus in adulthood (Boule et al., 2014). Consistent with its immunosuppressive effects, TCDD exposure suppresses the allergic immune response of rodents; this in turn results in decreased allergen-associated pathologic lung conditions and has been shown to suppress the development of experimental autoimmune disease (Quintana et al., 2008), to induce the suppression of autoimmune uveoretinitis (L. Zhang et al., 2010), and to affect colitis (Ji et al., 2015; Takamura et al., 2011), arthritis (Nakahama et al., 2011), and inflammatory lung diseases, such as silicosis (Beamer et al., 2012).
Experimental studies indicate that the AHR pathway plays an integral role in B-cell maturation, and that exposure to TCDD and other dioxin-like chemicals may alter B-cells and result in critical changes in the immune response (Baba et al., 2012; Sibilano et al., 2012; Simones and Shephard, 2011; Singh et al., 2011). Feng et al. (2016) found that chronic TCDD exposure impaired both B- and T-cell differentiation in a mouse model. Working with human B cells in vitro, Allan and Sherr (2010) demonstrated a new AHR-dependent mechanism by which exposure to environmental PAHs suppressed humoral immunity by blocking the differentiation of B cells into plasma cells. This finding was confirmed by data from human hematopoietic stem cells and knockout Ahr mouse models showing that the Ahr is critical in the maturation and differentiation of hematopoietic stem cells (Bock, 2017b; Fracchiolla et al., 2011; J. Li et al., 2017; Singh et al., 2011; Vaidyanathan et al., 2017). Furthermore, data from a B-cell specific Ahr knockout showed that the receptor pathway is required for efficient B-cell proliferation (Villa et al., 2017). Using a novel pluripotent stem cell–based culture system, B. W. Smith et al. (2013) demonstrated that AHR expression and activity can direct human hematopoietic progenitor cell proliferation and differentiation. These data show that pluripotent hematopoietic human cells express AHR and that AHR agonists enhance erythroid differentiation, whereas the antagonism of AHR favors the expansion of megakaryocyte cells. This finding supports previous work indicating that B-cell activation results in increased AHR expression and that an exposure of B-cells to benzo[a]pyrene, a PAH, suppresses B-cell differentiation (Allan and Sherr, 2010). H. Lu et al. (2010) demonstrated that although human B cells appeared less responsive to TCDD in increasing the expression of AHR battery genes, TCDD’s ability to decrease IgM production was similar in both mouse and human B cells. Data from Q. Zhang et al. (2013) suggest that this decrease in IgM production is the result of a TCDD-mediated decrease in B-cell
terminal differentiation, which results in fewer IgM-producing cells. Recent data from Kovalova et al. (2017) identify other species-specific gene expression in TCDD-exposed mouse, rat, and human B cells. In addition, Kovalova et al. (2016) found that certain AHR polymorphisms altered the sensitivity of human B cells to TCDD-mediated suppression of IgM secretion, consistent with AHR levels mediating this suppression.
TCDD not only alters hematopoietic stem cell maturation but also alters activation, proliferation, and migration in vivo and in vitro (Casado et al., 2011; Fader et al. 2015; Phadnis-Moghe et al., 2016), which indicates that exposure to it may have multiple effects on immune-cell function. Recent data have linked TCDD activation of the AHR with altered regulation of BCL-6 in human B-cells, which in turn is linked to non-Hodgkin lymphoma and diffuse large B-cell lymphomas (Phadnis-Moghe et al., 2015, 2016), providing a suggestive mechanistic link between TCDD exposure and some lymphomas.
Cellular immunity, which is mediated by the thymus and T cells, is also a target of TCDD and dioxin exposure and the AHR pathway (Baricza et al., 2016; Feng et al., 2016; Kuwatsuka et al., 2014). Early evidence indicated that dioxin and dioxin-like chemicals alter cellular immunity because it was observed that exposure to these chemicals resulted in thymic involution and suppressed cytotoxic T-lymphocyte activity (Hanieh, 2014). Recently attention has focused on the ability of the AHR to induce regulatory T cells, or Tregs (Bruhs et al., 2015; Kerkvliet, 2012; Marshall and Kerkvliet, 2010; Mohinta et al., 2015). A recent paper elucidates one potential molecular mechanism of AHR activation of Tregs, using real-time imaging to show the migration of AHR-Tr1 Tregs in the small intestine and the colon (Ehrlich et al., 2017). Tregs have potent suppressive activity in the immune system, and their inappropriate induction by TCDD could account for much of the immune suppression (Funatake et al., 2008; Kerkvliet, 2012; Marshall et al., 2008; Quintana et al., 2008; Stockinger et al., 2011; Yamamoto and Shlomchik, 2010). AHR activation in dendritic cells has also been shown to promote the development of Tregs by inducing tryptophan metabolism. Furthermore, a recent study shows a role for the AHR pathway in promoting gut Tregs (Ye et al., 2017).
One ultimate effect of the dysregulation of the immune system is an alteration in autoimmunity. Data from animal models and cell cultures indicate that exposure to dioxin and dioxin-like chemicals alters the development of autoimmune disorders. Boule et al. (2015) found that developmental exposure to TCDD exacerbated the severity of autoimmune disease in a genetically susceptible mouse model. In another example, antagonism of the AHR repressed the expression of cytokines and chemokines in primary human synovial fibroblasts (Lahoti et al., 2013), indicating a potential contribution to the inflammatory process of rheumatoid arthritis. N. T. Nguyen et al. (2013) hypothesized that the inflammatory process may occur when AHR stimulation of IL-17 production in Th17 cells overwhelms the immune suppressive effects of the inhibition of Treg
differentiation. TCDD has also been shown to induce apoptosis in rabbit chondrocytes, which supports a potential role of TCDD in contributing in a novel way to arthritis (Yang and Lee, 2010). A recent study indicates that sub-chronic low-dose TCDD exposure can show immunomodulatory effects in a mouse model of experimental autoimmune encephalitis (E. J. Yang et al., 2016). Recent work by L. Cheng et al. (2017) in a hospital-based study of Han Chinese patients suggests that a polymorphism in the AHR repressor gene may increase an individual’s susceptibility to rheumatoid arthritis. Exposure to TCDD was also shown to induce the reactivation of the latent form of the Epstein Barr virus in 19 patients with Sjögren syndrome, an autoimmune disease, when compared with 19 healthy patients (Inoue et al., 2012). AHR activation by ultraviolet light may play a role in the exacerbation of systemic lupus erythematosus symptoms by reducing DNA methylation in CD4+ T cells (Z. Wu et al., 2017). Taken together these studies suggest mechanisms by which TCDD may alter the incidence and progression of autoimmune disorders.
Allergies and allergic-induced asthma are also linked to TCDD exposure. A study of 18 people who had allergic asthma, 17 people whose asthma was controlled, and 12 controls showed that the plasma concentrations of IL-22 and the expression of the AHR in peripheral blood mononuclear cells were associated with the severity of allergic asthma; this finding strengthened the possibility that the AHR is involved in allergic asthma, thereby implying a role for dioxin exposure in this condition (Zhu et al., 2011). X. M. Li et al. (2016) studied a mouse model of non-allergic asthma and found that TCDD exposure reduced the airway infiltration of neutrophils and airway hyperresponsiveness and inhibited Th17 differentiation. Thus, depending on the disease, TCDD exposure could exacerbate or ameliorate symptoms.
Previous VAO committees have concluded that the data were inadequate or insufficient to support an increased risk of immune suppression, allergy, or autoimmune disease. The studies reviewed by these committees were at times poorly designed and often inconsistent and used a variety of biomarkers, making comparisons difficult. Most of the studies used biomarkers rather than health outcomes as endpoints.
The new studies reviewed here do not change this conclusion, as the results continue to be inconsistent and inconclusive. Cox et al. (2015) used hospital discharge records from New Zealand Vietnam veterans to examine the prevalence of inflammatory and autoimmune conditions. Although there was an increase in the standard hospitalization rate for rheumatoid arthritis but not systemic lupus erythematosus among veterans, no serum or tissue levels of dioxin-like chemicals were provided to confirm exposure. Rheumatoid arthritis incidence and prevalence were examined by Parks et al. (2016) in the female spouses of licensed
pesticide applicators in the AHS and found to be associated only with glyphosate, not 2,4-D or dicamba. Again, exposure was not confirmed with blood or tissue levels. Cappelletti et al. (2016) studied electric arc foundry workers exposed to the COIs. Results showed a statistically significant increase in rheumatoid arthritis among workers exposed to foundry dust. In a comparison of high- and low-exposure areas in Vietnam, C. H. Nguyen et al. (2017) showed that persons living in the high-exposure area had higher levels of AHR expression and some pro-inflammatory cytokines. They also had a higher prevalence of rheumatoid arthritis, but no data were provided linking the higher levels of pro-inflammatory cytokines in persons with rheumatoid arthritis. Among New Zealand workers in a plant that produced 2,4,5-T, comparisons of high- versus low-exposed workers by job and by serum measurements showed no difference in doctor-diagnosed nasal allergies, including hay fever.
It is biologically plausible for dioxins to influence immune dysfunction, but because of the variety of methods and endpoints used in studies to date, it is hard to confirm specific mechanisms by which the COIs induce immune suppression and auto-immunity. TCDD has been shown to suppress both limbs of the adaptive immune response, reduce the clearance of infection, and promote tumor growth (Bruhs et al., 2015; Kerkvliet, 2009, 2012; Kreitinger et al., 2016; Marshall and Kerkvliet, 2010). Exposure to dioxin-like chemicals has been shown to induce immune suppression via T regulatory cells (Bruhs et al., 2015; Kerkvliet, 2012; Marshall and Kerkvliet, 2010; Mohinta et al., 2015) in animal models. Animal models also support the development of inflammatory conditions, especially rheumatoid arthritis, following exposure to dioxin, through the AHR and downstream increased production of IL-17 (N. T. Nguyen et al., 2013). AHR polymorphisms have been associated with an increased susceptibility to rheumatoid arthritis in exposed persons (L. Cheng et al., 2017) and AHR activation by ultraviolet light may exacerbate systemic lupus erythematosus via reduced methylation in CD4+ T cells (Z. Wu et al., 2017). The data are inconsistent for allergic asthma.
On the basis of the evidence reviewed here and in previous VAO reports, the committee concludes that there is inadequate or insufficient evidence to determine whether there is an association between exposure to the COIs and any specific diseases involving immune suppression, allergy, autoimmunity, or inflammation.